Therapeutic methods and delivery systems utilizing sex steroid precursors

ABSTRACT

Sex steroid precursors such as dehydroepiandrosterone and dehydroepiandrosterone sulphate, and compounds converted in vivo to either of the foregoing, are utilized for the treatment of vaginal atrophy, hypogonadism, diminished libido, loss of collagen or connective tissues in the skin, and, in combination with an estrogen and/or progestin, for the treatment of menopause. The precursors may be formulated for percutaneous or transmucosal administration. Gels, solutions, lotions, creams, ointments and transdermal patches for the administration of these precursors are provided, as are certain pharmaceutical compositions and kits which can be used for the prevention and treatment of a wide variety of conditions related to decreased secretion of sex steroid precursors by the adrenals.

This is a division of application Ser. No. 08/005,619, filed Jan. 19,1993, now abandoned.

BACKGROUND OF THE INVENTION

This invention relates to a method for preventing and/or treatingvaginal atrophy, hypogonadism, diminished libido and menopause insusceptible warm-blooded animals including humans involvingadministration of dehydroepiandrosterone (DHEA),dehydroepiandrosterone-sulfate (DHEA-S) or compounds converted in vivoto either and to pharmaceutical products for delivery of activeingredient(s) useful to the invention.

Primates are unique in having adrenals that secrete large amounts of theprecursor steroid dehydroepiandrosterone (DHEA) and especiallyDHEA-sulfate (DHEA-S), which are converted into androstenedione(Δ4-dione) or androstene-diol (Δ⁵ -diol) and then into potent androgensand estrogens in peripheral tissues (Adams, Mol. Cell. Endocrinol. 41:1-17, 1985; Labrie et al., in Important Advances in Oncology (de VitaS., Hellman S., Rosenberg S. A., eds), J. B. Lippincott, PhiladelphiaPa., pp 193-200, 1985). DHEA-S, the major steroid present in blood ofboth men and women is converted into DHEA and Δ5-diol in peripheraltissues, thus maintaining a close correlation between the concentrationof these three steroids in the blood (Adams, Mol. Cell. Endocrinol. 41:1-17, 1985). Depending upon the relative activities of17β-hydroxysteroid dehydrogenase (17β-HSD), aromarase and 5α-reductase,DHEA or its derivatives will be preferentially converted into androgensand/or estrogens.

The low serum values of DHEA and DHEA-S found at birth persist up to sixyears of age. Usually, during the 7th year of age, serum levels of thesetwo steroids increase and continue to rise until age 16 in both boys andgirls (Orentreich et al., J. Clin. Endocr. Metab. 59: 551-555, 1984). Afurther increase is then seen in males, who typically reach maximallevels between 20 and 24 years of age. In women, there is usually nofurther increase after 16 years. DHEA and DHEA-S decrease with aging inboth men and women (Vermeulen and Verdoreck, J. Steroid Biochem. 7:1-10, 1976; Vermeulen et al., J. Clin. Endocr. Metab. 54: 187-191,1982). In fact, at 70 years of age, serum DHEA-S levels are atapproximately 20% of their peak values while they decrease by up to 95%by the age of 85 to 90 years (Migeon et al., J. Clin. Endocr. Metab. 17:1051-1062, 1957). The 70% to 95% reduction in the formation of DHEA-S bythe adrenals during aging results in a dramatic reduction in theformation of androgens and estrogens in peripheral target tissues, thusresulting in a marked decrease in the biochemical and cellular functionsinduced by sex steroids.

In addition to the decrease in DHEA-S formation, in men, a progressivedecrease in the concentration of testosterone in the spermatic vein(Giusti et al., Exp. Gerontol. 10: 241-245, 1975) as well as in plasma(Lewis et al., Acta Endocrinol. 82: 444-448, 1976; Zumoff et al., J.Clin. Endocr. Metab. 54: 534-538, 1982) has been observed after the ageof 60-70 years. Such data, however, have been the subject of controversy(Nieschlag et al., J. Clin. Endocr. Metab. 55: 676-681, 1982). Inpostmenopausal women, serum testosterone levels are lower than duringreproductive life (Forest M. G., Physiological changes in circulatingandrogens, in Androgens in Childhood (Forest M. G., ed), Karger Basel,p. 104-129, 1989).

The skin is an important site of sex steroid formation and its functionis known to be regulated by sex steroids. Sex steroids can act directlyin the skin or may stimulate growth hormone and prolactin secretion bythe anterior pituitary gland. Skin atrophy is in fact known to occur ingrowth hormone defidency, probably through a secondary decrease ininsulin-like growth factor (IGF-1) secretion. Serum growth hormone (GH)and insulin-like growth factor (IGF-1) levels are known to be reduced inaging men and women.

Plasma DHEA-S concentration has been suggested as a predictor ofosteoporosis (Nordin et al., J. Clin. Endocr. Metab. 60: 651-657, 1985;Deutsch et al., Int. J. Gynecol. Obstet. 25: 217-220, 1987). Serum DHEAhas in fact been found to be significantly lower in osteoporoticcompared to normal subjects (Nordin et al., J. Clin. Endocr. Metab. 60:651-657, 1985). Low tissue Δ4-dione is also likely to accompany lowDHEA-S levels. Since Δ4-dione is the precursor of estrone which is amain source of estradiol in postmenopausal women (Marshall et al., Clin.Endocrinol. 9: 407, 1978), such secondary low levels of estrogens arelikely to be involved in osteoporosis (Nordin et al., Lancet 2: 277,1981).

As another mechanism, low serum DHEA-S levels resulting in low formationof androgens in peripheral tissues could also result in low boneformation, a characteristic of postmenopausal osteoporosis (Meunier etal., in Histological heterogeneity of apparently idiopathic osteoporosisand treatment (DeLuca H. F., Frost H. M., Jee W. S. S., Johnston Jr C.C., Parfitt A. M., eds), University Park Press, Baltimore, p. 293). Infact, Deutsch et al. (Int. J. Gynecol. Obstet. 25: 217-222, 1987) havefound a significant correlation between serum DHEA-S and androstenedionelevels and osteoporosis in late postmenopausal women while nocorrelation was found between serum estrogens and bone density, thussuggesting a major importance of androgens in postmenopausal bone loss.

Studies in animals have shown that androgen deficiency leads toosteopenia while testosterone administration increases the overallquantity of bone (Silverberg and Silverberg, 1971; See Finkelstein etal., Ann. Int. Med. 106: 354-361, 1987). Orchiectomy in rats can causeosteoporosis detectable within 2 months (Winks and Felts, Calcif.Tissue. Res. 32: 77-82, 1980; Verhas et al., Calcif. Tissue Res. 39:74-77, 1986).

As mentioned earlier, adrenal androgen levels have been found to bereduced in osteoporosis (Nordin et al., J. Clin. Endocr. Metab. 60: 651,1985). Moreover, elevated androgens in postmenopausal women have beenshown to protect against accelerated bone loss (Deutsch et al., Int. J.Gynecol. Obstet. 25: 217-222, 1987; Aloia et al., Arch. Int. Med. 143:1700-1704, 1983). In agreement with such a role of androgens, urinarylevels of androgen metabolites are lower in postmenopausal symptomaticmenopausis than in matched controls and a significant decrease inconjugated dehydroepiandrosterone (DHEA) was found in the plasma ofosteoporotic patients (Hollo and Feher, Acta Med. Hung. 20: 133, 1964;Urist and Vincent, J. Clin. Orthop. 18: 199, 1961; Hollo et al., ActaMed. Hung. 27: 155, 1970). It has been suggested that postmenopausalosteoporosis results from both hypoestrogenism and hypoandrogenism(Hollo et al., Lancet, 1357, 1976).

As a mechanism for the above-suggested role of both estrogens andandrogens in osteoporosis, the presence of estrogen (Komm et al.,Science 241: 81-84, 1988; Eriksen et al., Science 241: 84-86, 1988) aswell as androgen (Colvard et al., Proc. Natl. Acad. Sci. 86: 854-857,1989) receptors in osteoblasts could explain increased bone resorptionobserved after estrogen and androgen depletion.

While, in women, there is a rapid bone loss starting at menopause, boneloss in males can be recognized at about 65 years of age (Riggs et al.,J. Clin. Invest. 67: 328-335, 1987). A more significant bone loss isseen in men at about 80 years of age, with the accompanying occurrenceof hip, spine and wrist fractures. Several studies indicate thatosteoporosis is a clinical manifestation of androgen deficiency in men(Baran et al., Calcif. Tissue Res. 26: 103-106, 1978; Odell andSwerdloff, West J. Med. 124: 446-475, 1976; Smith and Walker, Calif.Tissue Res. 22 (Suppl.): 225-228, 1976).

Therapy of postmenopausal women with nandrolone increased cortical bonemineral content (Clin. Orthop. 225: 273-277). Androgenic side-effects,however, were recorded in 50% of patients. Such data are of interestsince while most therapies are limited to an arrest of bone loss, anincrease in bone mass was found with the use of the anabolic steroidnandrolone. A similar stimulation of bone formation by androgens hasbeen suggested in a hypogonadal male (Baran et al., Calcif. Tissue Res.26: 103, 1978).

The decline with age of serum levels of DHEA-S and DHEA has led to theintriguing possibility that low serum DHEA and DHEA-S levels could beassociated with cancer and cardiovascular diseases. In fact, a series ofstudies have indicated that subnormal levels of DHEA are associated witha high risk of breast cancer (Bulbrook et al., Lancet 2: 395-398, 1971;Rose et al., Eur. J. Cancer 13: 43-47, 1977; Thijssen et al., J. SteroidBiochem. 6: 729-734, 1975; Wang et al., Eur. J. Cancer 10: 477-482,1974; Gomes et al., C. R. Acad. Sci. Paris 306: 261-264, 1978; Brownsezet al., Eur. J. Cancer 8: 131-137, 1972). Women with breast cancer werefound to have low urinary levels of androsterone and etiocholanolone,two metabolites of DHEA (Bulbrook et al., Lancet 2:1238-1240, 1962;Cameron et al., Br. Med. J. 4: 768-771, 1970). Bulbrook et al. (Lancet2: 1235-1240, 1962) then reported that women with primary operablebreast cancer had urinary levels of 11-deoxy-17-ketosteroids (derivedmainly from DHEA-S and DHEA) lower than normal, thus suggesting that alow secretion rate of DHEA and DHEA-S could precede the development ofbreast cancer.

The main approaches for the treatment of already developed breast cancerare related to the inhibition of estrogen action and/or formation. Therole of estrogens in promoting the growth of estrogen-sensitive breastcancer has been recognized (Lippman, Semin. Oncol. 10 (Suppl. 4): 11-19,1983; Sledge and McGuire, Cancer Res. 38: 61-75, 1984; Witliff, Cancer53: 630-643, 1984; Poulin and Labrie, Cancer Res. 46: 4933-4937, 1986).

DHEA (450 mg/kg, b.w., 3 times a week) markedly delayed the appearanceof breast tumors in C3H mice which were genetically bred to developbreast cancer (Schwartz, Cancer Res. 39: 1129-1132, 1979). Moreover, therisk of developing bladder cancer was found to be increased in menhaving lower serum DHEA levels (Gordon et al., Cancer Res. 51:1366-1369, 1991).

Copending U.S. patent application Ser. No. 07/785,890 filed Nov. 4,1991, relates to a method of treatment of breast and endometrial cancerin susceptible warm-blooded animals which may include inhibition ofovarian hormonal secretion by surgical means (ovariectomy) or chemicalmeans (use of an LHRH agonist, e.g. D-Trp⁶, des-Gly-NH₂ ¹⁰ !LHRHethylamide, or antagonist) as part of a combination therapy.Antiestrogens, androgens, progestins, inhibitors of sex steroidformation (especially of 17β-hydroxysteroid dehydrogenase- oraromarase-catalyzed production of sex steroids), inhibitors of prolactinsecretion and of growth hormone secretion and ACTH secretion arediscussed.

Copending U.S. patent application Ser. No. 07/724,532 filed on Jun. 28,1991, relates to a method using low dose androgenic compounds for theprevention and treatment of breast cancer, endometrial cancer,osteoporosis and endometriosis.

Recent in vitro studies describe the relative antiproliferativeactivities of an androgen on the growth of the estrogen-sensitive humanmammary carcinoma cell line ZR-75-1 (Poulin et al. "Androgens inhibitbasal and estrogen-induced cell proliferation in the ZR-75-1 humanbreast cancer cell line", Breast Cancer Res. Treatm. 12: 213-225, 1989).As mentioned above, Poulin et al. (Breast Cancer Res. Treatm. 12:213-225, 1989) have found that the growth of ZR-75-1 human breastcarcinoma cells is inhibited by androgens, the inhibitory effect ofandrogens being additive to that of an antiestrogen. The inhibitoryeffect of androgens on the growth of human breast carcinoma cellsZR-75-1 has also been observed in vivo in nude mice (Dauvois and Labrie,Cancer Res. 51: 3131-3135, 1991).

DHEA has been suggested to have beneficial effects in obesity, diabetes,atherosclerosis, cancer, autoimmune diseases, fatigue, loss of musclemass, connective tissue diseases, aging and longevity (Orentreich etal., J. Clin. Endocrinol. Metab. 59: 551-555, 1984; Regelson, Ann. N.Y.Acad. Sci. 521: 260-273, 1988; Gordon et al., Adv. Enzyme Regul. 26:355-383, 1987; Schwartz, Adv. Cancer Res. 51: 391-423, 1988;Barrett-Connor et al., New Engl. J. Med. 315: 1519-1524, 1986).

In aged Sprague-Dawley rats, Schwartz (in Kent, Geriatrics 37: 157-160,1982) has observed that body weight was reduced from 600 to 550 g byDHEA without affecting food intake. Schwartz (Cancer 39: 1129-1132,1979) observed that C3H mice given DHEA (450 mg/kg, 3 times a week)gained significantly less weight and grew older than the controlanimals, had less body fat and were more active. The reduction in bodyweight was achieved without loss of appetite or food restriction.Furthermore, DHEA could prevent weight gain in animals bred to becomeobese in adulthood (in Kent, Geriatrics 37: 157-160, 1982).

DHEA in the diet has been shown to be a potent antihyperglycemic andantidiabetic agent in mice with inherited obesity-glucose intolerancesyndrome (Coleman et al., Diabetes 33: 26-32, 1984).

DHEA reduced the incidence of atherosclerosis in cholesterol-fed rabbits(Gordon et al., J. Clin. Invest. 82: 712-720, 1988; Arad et al.,Arteriosclerosis 9: 159-166, 1989). Moreover, high serum concentrationsof DHEA-S have been reported to protect against death fromcardiovascular diseases in men (Barrett-Connor et al., N. Engl. J. Med.315: 1519-1524, 1986). Circulating levels of DHEA and DHEA-S have thusbeen found to be inversely correlated with mortality from cardiovasculardisease (Barret-Connor et al., N. Engl. J. Med. 315: 1519-1524, 1986)and to decrease in parallel with the diminished immune competence(Thoman and Weigle, Adv. Immunol. 46: 221-222, 1989). A study in man hasshown an inverse correlation between fetal serum DHEA-S and low densitylipoprotein (LDL) levels (Parker et al., Science 208: 512, 1980).

In normal men, a placebo-controlled trial investigated the effect ofdaily oral administration of 1.6 g of DHEA for 28 days. Serum DHEAlevels were increased 2.5- to 3.5-fold in the DHEA-treated group whiletotal cholesterol and serum LDL cholesterol decreased by 7.1 and 7.5%,respectively (Nestler et al., J. Clin. Endocrinol. Metab. 66: 57-61,1988). A decrease in body fat was found in 4 of the 5 men treated withDHEA for an average 31% decrease in percent of body fat with no changein total weight, thus suggesting a corresponding increase in musclemass.

Drucker et al. (J. Clin. Endocrinol. Metab. 35, 48, 1972), Buster et al.(Am. J. Obstet. Gynecol. 166, 1163, 1992) and Welle et al. (J. Clin.Endocrinol. Metab., 71, 1259, 1990) have also administered DHEA orallyto the human.

Obesity was found to be improved in the A^(vy) mutant mouse (Yen et al.,Lipids 12: 409-413, 1977) and in the Zucker rat (Cleary and Zisk, Fed.Proc. 42: 536, 1983). DHEA-treated C3H mice had a younger appearancethan controls (Schwartz, Cancer Res. 39: 1129-1132, 1979).

Brain concentrations of DHEA are 6.5 times higher than correspondingconcentrations in plasma (Lacroix et al., J. Steroid Biochem. 28:317-325, 987). DHEA and DHEA-S improve memory in aging mice (Flood andRoberts, Brain Res. 448: 178-181, 1988). Serum DHEA-S concentrations inpatients with Alzheimer disease have been found to be 48% lower onaverage than in age-matched controls (Sunderland et al., Lancet ii: 570,1989). As mentioned above, DHEA administered chronically in the diet hasbeen shown to increase longevity by delaying the development of somediseases in particular strains of animals.

U.S. Pat. No. 4,496,556 describes the use of DHEA or its derivatives totreat skin dryness by topical administration. Only local action on thesebaceous glands was described and no systemic action was observed.

U.S. Pat. No. 4,542,129 describes a topical composition for treating dryskin in a patient comprising the combination of DHEA and/or derivatives,a keratolytic agent and a non-toxic dermatologically acceptable vehicle.

Great Britain Patent No 1246639 describes preparation of esters ofdehydroepiandrosterone for use as agents in the treatment of post andpremenopause, tachycardia and headaches.

One problem facing the use of DHEA in humans is that high doses arerequired apparently because a large proportion of the compound isdegraded in the liver before it reaches the blood stream after oraladministration.

It is known that the efficiency of delivery of some drug can be improvedby the use of certain pharmacologically inactive derivatives which are,by in vivo enzymatic or spontaneous reactions, transformed into theactive drugs (see generally H. Bundgaard, Design and application ofprodrugs. In A textbook of Drug Design and Development. Edited by P.Krogsgaard-Larsen and H. Bundgaard. Harwood Academic Publishers GmfH,Chur, Switzerland, 1991, pp. 113-191). For example, Druzgala et al., J.Steroid Biochem. Molec. Biol. 38, 149-154, 1991, describes prodrugs ofglucocorticoids. Bodor et al. in U.S. patent application Ser. No.4,213,978 and in German Patent Application Publication No DE 29 48 733disclose the use of thiazolidine derivatives of progesterone as topicaldrugs. Percutaneous absorption of prodrug derivatives of estrogens andprogestins are reported by Friend D. R. in Critical Reviews inTherapeutic Drug Carrier Systems, vol. 7 (2), pp. 149-186, 1990.Information about percutaneous absorption of drugs can also be found inSkin Permeability (H. Schaefer, A. Zesch and G. Stuttgen, eds),Springer-Verlag, Berlin, Heidelberg, New York, 1982, pp. 896.

Currently, low dose estrogen therapy is the standard approach used inperimenopausal and menopausal women to relieve vasomotor symptoms,urogenital atrophy, osteoporosis and other symptoms and signs associatedwith menopause (for review, see Edman, C. D., Estrogen ReplacementTherapy. In: The menopause, Springer-Verlag, New York, (edited by H. J.Buchsbaum), pp. 77-84, 1983). Detailed information about the menopauseand its therapy can be found in other chapters of this book. Obviously,such approaches limited to estrogen replacement therapy, associated ornot with progestins, do not reproduce the equilibrium between estrogensand androgens that occurs naturally from the transformation of DHEA intoits active metabolites in various target tissues.

Some esters of DHEA at position 3 are already described in thelitterature (Riva et al., J. Org. Chem. 54: 3161-4, 1989; Parish andChistrakorn, Synth. Commun. 15: 393-9, 1985; Rom Patent No RO 66924B;Jarosz and Zamojski, Tetrahedron 38:1453-6, 1982; Heublin et al., Z.Chem. 22: 178, 1982; German Patent Application No DE 2534911; Khaidem etal., Indian J. Chem. Sect. B, 27B: 850-1, 1988; Petit et al., J. Org.Chem. 52: 3573-8, 1987; Hanson and Reese, J. Chem. Soc. Perkin Trans. 1:647-649, 1985); European Patent Application No 84-105741; Heublein etal., Acta Polym., 35: 673-7, 1984; Seevers et al., J. Med. Chem., 25:1500-3, 1982; Yamashita and Kurosawa, Agric. Biol. Chem., 39: 2243-4,1975; Japan Patent Application JP 50005372; Pohlmann et al., Mol. Cryst.Liquid Cryst. 13: 243-54, 1971.

Alkanesulfonates of DHEA is described as inhibitors ofglucose-6-phosphate dehydrogenase activity in J. Pharm. Sci. 73: 1643-5,1984.

In Britain Patent Application No GB 1246659 and S. Africa PatentApplication No ZA 6806112 is disclosed DHEA esters for the treatmentrespectively of post and premenopause tachycardia and headaches andclimateric complaints.

Leszczynski et al., in Biochem. Biophys. Acta, 1014: 90-7, 1989; idem:1083: 18-28, 1991, have reported esterification of DHEA by blood plasmaand Katz et al., in MCF-7 cell line in J. Steroid Biochem, 26: 687-92,1987.

Ethyl carbonate of DHEA is reported by Weisz and Agocs in Arch. Pharm.(Weinheim, Ger), 319: 952-3, 1986.

Some halogeno esters of DHEA are described by Challis and Heap in J.Chromatogr. 50: 228-238, 1970 and by Pinelly and Nair in J. Chromatogr.43: 223-228, 1969.

SUMMARY OF THE INVENTION

It is an object of the present invention to provide simple and efficientmethods for systemic delivery of sex steroid precursors such asdehydroepiandiosterone, dehydroepiandrosterone sulfate and analogs whichare converted in vivo to either DHEA or DHEA-S.

It is another object of the invention to provide novel treatments ofmenopause, vaginal atrophy, hypogonadism and diminished libido.

It is another object of the invention to provide kits and pharmaceuticalcompositions for use in accordance with the invention.

It is another object of the invention to provide novel sex steroidprecursors and pharmaceutical compositions thereof.

In one aspect, the invention provides a method for treating menopausecomprising administering to a patient in need of such treatment aneffective amount of at least one sex steroid precursor selected from thegroup consisting of dehydroepiandrosterone, dehydroepiandrosteronesulphate, and compounds converted in vivo to either of the foregoing, incombination with an effective amount of an estrogen, a progestin orboth.

In another aspect, the invention provides a pharmaceutical compositionfor the treatment of menopause comprising at least one sex steroidprecursor selected from the group consisting of dehydroepiandrosterone,dehydroepiandrosterone sulphate, and compounds converted in vivo toeither of the foregoing, and further comprising an estrogen or aprogestin or both. One preferred combination is precursor and estrogen.

In another aspect, the invention provides a kit for the treatment ofmenopause having a first container which includes at least one sexsteroid precursor selected from the group consisting ofdehydroepiandrosterone, dehydroepiandrosterone sulphate, and compoundsconverted in vivo to either of the foregoing, and at least oneadditional container having either a progestin, an estrogen, or both.One preferred kit includes three containers having a precursor, anestrogen and a progestin, respectively, in separate containers.

In another aspect, the invention provides a method of treating vaginalatrophy, hypogonadism, diminished libido or reduced collagen orconnective tissue in the skin comprising administering to a patient inneed of such treatment an effective amount of a sex steroid precursorselected from the group consisting of dehydroepiandrosterone,dehydroepiandrosterone sulphate, and compounds converted in vivo toeither of the foregoing.

In another aspect, the invention provides a therapeutic method oftreating reduced or imbalanced concentrations of sex steroids comprisingapplying an effective amount of a pharmaceutical composition forpercutaneous or transmucosal delivery to an outer surface of skin ormucosa of a patient in need of such treatment, said pharmaceuticalcomposition comprising a carrier having dissolved therein at least onesex steroid precursor selected from the group consisting ofdehydroepiandrosterone, dehydroepiandrosterone sulphate, and compoundsconverted in vivo to either of the foregoing, said precursor beingpresent at a concentration of at least 7% by weight relative to thetotal pharmaceutical composition, and said carrier being compatible withskin or mucosa and allowing penetration of said precursor through saidskin or mucosa, said carrier having sufficient viscosity to maintainsaid precursor on a localized area of skin or mucosa, without running orevaporating, for a time period sufficient to permit substantialpenetration of said precursor through said localized area of said skinor mucosa. The foregoing method is useful in treating the conditionsdiscussed above, menopausal symptoms and other conditions which respondto replenishment of diminished DHEA levels, including but not limited toobesity, cardiovascular disease, atherosclerosis, breast cancer,endometrial cancer, loss of muscle mass, diabetes, fatigue, connectivetissue diseases and memory loss.

In another aspect, the invention provides a pharmaceutical compositionfor percutaneous or transmucosal delivery of said pharmaceuticalcomposition comprising a carrier having dissolved therein at least onesex steroid precursor selected from the group consisting ofdehydroepiandrosterone, dehydroepiandrosterone sulphate, and compoundsconverted in vivo to either of the foregoing, said precursor beingpresent at a concentration of at least 7% by weight relative to thetotal pharmaceutical composition, and said carrier being compatible withskin or mucosa and allowing penetration of said precursor through saidskin or mucosa, said carrier having sufficient viscosity to maintainsaid precursor on a localized area of skin or mucosa, without running orevaporating, for a time period sufficient to permit substantialpenetration of said precursor through said localized area of said skinor mucosa.

In another aspect, the invention provides novel compounds (andpharmaceutical compositions containing them) of the formulas set forthbelow with substituent definitions set forth below: ##STR1## wherein Xis selected from the group consisting of H, ##STR2## RCO₂ CHR^(a) -- andR^(b) SO₂ --

R being selected from the group consisting of hydrogen, straight- orbranched-alkyl, straight- or branched-alkenyl, straight- orbranched-alkynyl, aryl, furyl, straight- or branched-alkoxy, straight-or branched-alkenyloxy, straight- or branched-alkynyloxy, aryloxy,furyloxy and halogeno analogs of the foregoing

R^(a) being hydrogen or (C₁ -C₆) alkyl; and

R^(b) being selected from the group consisting of hydroxyl (or saltsthereof), methyl, phenyl and p-toluyl;

wherein Y is a divalent ;substituted or unsubstituted moiety of theformula: ##STR3##

(Z being an oxygen or sulfur atom), and wherein Y and the carbon atom towhich it is bonded together form a closed saturated 5-membered ring,##STR4## wherein R^(c) is selected from the group consisting of C₃ -C₂₀straight- or branched-alkyl, C₃ -C₂₀ straight- or branched-alkenyl, C₃-C₂₀ straight- or branched-alkynyl, aryl and halo-substituted analogs ofthe foregoing. ##STR5##

R^(d) being selected from the group consisting of hydrogen, straight- orbranched-alkyl, straight- or branched-alkenyl, straight- orbranched-alkynyl, aryl, furyl, straight- or branched-alkoxy, straight-or branched-alkenyloxy, straight- or branched-alkynyloxy, aryloxy,furyloxy and halogeno analogs of the foregoing.

wherein R_(a) is hydrogen or (C₁ -C₆) alkyl. ##STR6## wherein R³ ishydroxy or sulfate. wherein R^(e) is selected from the group consistingof hydrogen, benzyl, aryl, straight- or branched-alkyl, straight- orbranched-alkenyl and straight- or branched-alkynyl.

wherein Z is oxygen or sulfur.

In one embodiment, a method is provided to compensate for the markeddecrease in the secretion of the sex steroid precursors DHEA and DHEA-Sby the adrenals during aging comprising administering DHEA, DHEA-S oranalogs converted in vivo thereto in amounts which compensate for theconsequences of decreased DHEA and DHEA-S secretion by the agingadrenals without exerting unwanted side effects.

It is believed that the methods of the invention are suitable for bothprophylactic and therapeutic use. The serum concentrations, kits, andcompositions discussed herein are equally useful toward eitherobjective.

In another aspect, the invention provides a transdermal devicecomprising;

(a) a surface adapted for contact with human skin;

(b) a means of maintaining said surface on a localized area of skin towhich said device is applied;

(c) a storage member in fluid communication with said surface, saidmember containing a pharmaceutical composition comprising a carrier andan active ingredient selected from the group consisting ofdehydroepiandrosterone, dehydroepiandrosterone sulphate and analogueswhich are converted in vivo to either; and

(d) a means for conducting said pharmaceutical composition from saidstorage member through said surface and into contact with said localizedarea of skin.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

To select patients who may benefit from the treatments described herein,the serum levels of DHEA and its metabolites can be measured asdescribed by Belanger et al., in Steroid Formation, Degradation andAction in Peripheral, Normal and Neoplastic Tissues (H. Bradlow, L.Castagnetta, S. d'Aquino, L. Gogliotti, eds) Ann. N.Y. Acad. Sci. 586:93-100, 1990; Haning et al., J. Clin. Endocrinol. Metab. 72: 1088, 1991.See also Labrie et al., Endocrinology 123, 1412-1417, 1988. Serum IGF-1levels can be measured as described (Furlanetto et al., J. Clin. Invest.60: 648, 1977). In accordance with the invention, once the DHEAdificiency is determined, DHEA or its analogues is administered at adosage sufficient to cause and maintain serum DHEA concentration between4 and 7 micrograms per liter. When DHEA is administered by the preferredpercutaneous or transmucosal technique, it has been found that DHEA isvery efficiently absorbed into the blood to raise serum levels. Forexample, when a Glaxal cream, containing Glaxal base as a carrieravailable from Glaxal Canada Limited and 10% DHEA by weight of the totalcomposition, is applied twice daily to a 100 square centimeter surfaceof the abdominal area in an amount providing 100 milligrams of activeingredient (e.g. DHEA), a typical patient is likely to respond with anincrease of serum DHEA concentration of about 0.7 micrograms per literper 50 kg of body weight. The delivered dosage may be raised or loweredin known manners by altering the location to which the lotion orointment is applied, by altering the size of the surface area to whichit is supplied, by altering the concentration of the active ingredient,or by altering the carrier. For example, increasing the surface areawill normally increase the dosage of active ingredient delivered if theconcentration of active ingredient remains constant. In the same manner,dosage delivered increases with increased concentration of activeingredient in the delivery base, and decreases with decreasedconcentration. Dosage delivered into the bloodstream also varies in aknown manner with respect to the body region at which the transdermalpenetration system is applied to the skin. Changing the carrier can alsoalter the delivered dosage in known ways. Preferably, serum DHEAconcentration is measured before treatment begins, and a dosage isselected to quickly raise serum DHEA concentration to the preferredtarget range between 4 and 7 micrograms per liter. Subsequently, thepatient is monitored both symptomatologically and by DHEA concentrationto verify that the desired serum concentration target and symptomaticrelief have been obtained. DHEA is then maintained a a constantconcentration in the circulation. For a typical post-menopausal patient,for example, this dosage is the equivalent of application of 400 mg ofthe active precursor, as part of a 10 percent composition in Glaxal, toa 400 square centimeter area of the abdomen 2 times daily per 50 kg ofbody weight. If oral administration is chosen, 800 mg should beadministered twice daily per 50 kg of body weight

In accordance with the invention, DHEA, DHEA-S and/or compoundsconverted to either in vivo are utilized for the treatment of menopause,vaginal atrophy, hypogonadism, or diminished libido. Additionally, otherconditions related to decreased secretion of DHEA by the adrenals duringaging and which respond to DHEA therapy can be treated more efficientlywith transdermally delivered DHEA, DHEA-S (or analogues) in accordancewith the invention. Conditions expected to respond to the treatmentsherein may be diagnosed in conventional ways. For example, theappearance of breast cancer is usually detected by self breastexamination, clinical breast examination by the physician and/ormammography. Endometrial cancer, on the other hand, is usually diagnosedby the PAP smear and/or endometrial biopsy. Both cancers can bediagnosed and evaluated by standard physical methods well known to thoseskilled in the art, e.g. bone scan, chest X-Ray, skeletal survey,ultrasonography of the liver and liver scan (if needed), CAT scan, MRIand physical examination.

The first manifestations of menopause are usually hot flashes. Furthercharacterization of menopause can be determined in accordance with knowntechniques. See for Example, The Menopause (Herbert J., Buchsbaurm, ed),Springer Verlag, New York (983), pp. 222.

Vaginal atrophy is often indicated by dyspareunia and vaginalinfections. Vaginal atrophy, hypogonadism and diminished libido are allcharacterized in well-known ways. For the above-indicated diseases, see,for example, Korenman, Stanley G., "Sexual Dysfunctions" in WilliamsTextbook of Endocrinology (Jean D. Wilson and Daniel W. Foster, Eds.),W. B. Saunders Co, Philadelphia, pp. 1033-1048, 1992.

Bone density, on the other hand, can be measured by standard methodswell known to those skilled in the art, e.g. QDR (Quantitative DigitalRadiography), dual photon absorptiometry and computerized tomography.Plasma and urinary calcium and phosphate levels, plasma alkalinephosphatase, calcitonin and parathormone concentrations, as well asurinary hydroxyproline and calcium/creatinine ratios are usefulparameters of bone formation and resorption.

Loss of collagen or connective tissues in the skin often accompaniesaging, espedally in persons over 50 years of age. It may be evidenced bywrinkling of the skin and/or low elasticity.

Osteoporosis or otherwise insufficient bone mass, and other diseasestreatable by activating the androgen receptor may be treated inaccordance with the present invention or prophylactically prevented inaccordance herewith. The present invention can aid in the prevention ofbreast or endometrial cancer.

The normal range of body weight is well known to those skilled in theart, while cholesterol and lipoproteins are routinely measured bystandard techniques (Nestler et al. J. Clin. Endocrinol. Metab. 66:57-61, 1988 for references).

Skin status can be assessed by visual inspection, palpation and, withmore precision, by punch biopsy and standard histological examination.

In one preferred treatment for menopause, the invention seeks tosimultaneously maintain blood levels of estrogen and a sex steroidprecursor (e.g., DHEA or DHEA-S) within normal pre-menopausalparameters. The body converts DHEA-S to DHEA in most peripheral tissues.Without intending to be bound by theory, it is believed that maintenanceof appropriate precursor levels will better enable natural enzymes, suchas 17β-hydroxysteroid dehydrogenase, 3β-hydroxysteroidhydrogenase,aromarase and 5α-reductase to regulate production of androgens andestrogens and to maintain them in a manner more closely resembling theirrelative levels prevailing prior to menopause. Hence, the inventioncontemplates that not only estrogens but also androgens and precursorswill be kept in better balance. In fact, all target tissues possess theenzymatic machinery necessary to synthesize androgen and/or estrogensaccording to local control and need (Labrie, Mol. Cell. Endocrinol. 78,C113-C118, 1991).

The estrogen and precursor (e.g., DHEA) when administered together inaccordance with the invention may be administered simultaneously orseparately. Indeed, the second active ingedient may be added to anexisting estrogen therapy in order to achieve the combination therapy ofthe invention.

It is necessary only that both the estrogen and precursor beadministered in a manner and at a dosage sufficient to allow blood serumconcentration of each to obtain desired levels. In accordance with thecombination therapy of the invention, concentration of the precursor ismaintained within desired parameters at the same time that estrogenconcentration is maintained within desired parameters. Where estradiolis used, serum estradiol concentration should typically be maintainedbetween 50 and 300 nanograms per liter, preferably between 100 and 200nanograms per liter and most preferably between 150 and 175 nanogramsper liter. Where another estrogen is used, serum concentration may bevaried in a known manner to account for the difference in estrogenicactivity relative to estradiol and in order to achieve normalpre-menopausal estrogen levels. A lesser concentration is needed, forexample, if Mestranol is used. Adequate serum estrogen levels can alsobe assessed by disappearance of the symptoms of menopause. Serumconcentration of the second compound of the combination therapy (e.g.,DHEA) is typically maintained between 4 and 7 micrograms per liter,preferably between 5 and 7 micrograms per liter, and most preferablybetween 6 and 7 micrograms per liter.

The estrogen is preferably estradiol, but may be sodium estrone sulfateor another compound which acts as an estrogen receptor agonist. Whenadministered separately, commercially available estrogen supplements maybe used, e.g., PREMARIN available from Ayerst (St-Laurent, Quebec,Canada). One preferred precursor is DHEA, although DHEA-S and analogsdiscussed below are also especially effective for the reasons statedbelow. Where DHEA is used, for example, DHEA, pharmaceutical grade, iscommercialy available from SIGMA (St-Louis, Mo., U.S.A.). For typicalpatients, the appropriate dosage of estrogen to achieve desired serumconcentrations is between 0.3 and 2.5 milligrams of PREMARIN per day per50 kg of body weight when administered orally. In certain embodiments ofthe invention, the estrogen may be 17β-estradiol administeredpercutaneously in a patch which is available from CIBA under the nameESTRADERM wherein the daily dose is between 0.05 and 0.2 milligrams perday per 50 kg of body weight. For typical patients, the appropriatedosage of the sex steroid precursor to achieve desired serumconcentration of the precursor (or its metabolite where the precursor isconverted in vivo to DHEA or DHEA-S) is between 0.25 and 2.5 grams perday per 50 kg of body weight when administered orally. The precursor mayalso be administered transdermally, as described in more detail below,in a sufficient amount to achieve target serum concentration. Thatcorrelation is also discussed in more detail below.

In another embodiment, menopause is treated with precursor as set forthabove, in combination with periodic administration of a progestin suchas medroxyprogesterone acetate (e.g. Provera) which is preferablyadministered intermittently, e.g. at a dosage of 5-10 mg per day for 10consecutive days, said 10-day periods being spaced 20 days to 5 months apart. A combination therapy using a precursor, an estrogen and aprogestin may also be used, preferably at the dosages discussed abovefor each component.

The sex steroid precursor used in the invention may be administered withor without additional carrier or diluent by the oral route but requiresan additional carrier or diluent when administered by the preferredpercutaneous or transmucosal route. In a pharmaceutical composition fororal administration, DHEA or other precursor is preferably present in aconcentration between 5 and 98% by weight relative to total weight ofthe composition more preferably between 50 and 98 percent, especiallybetween 80 and 98 percent. If estrogen such as estradiol is present, itsconcentration is preferably from 0.04 to 0.4 percent by weight. A singleprecursor such as DHEA may be the only active ingredient, oralternatively, a plurality of precursors and/or their analogues may beused (e.g., a combination of DHEA and DHEA-S, or a combination of two ormore compounds converted in vivo to DHEA or DHEA-S, or a combination ofDHEA and one or more analogues thereof which are converted to DHEA invivo, etc. Where a combination is used, the total dosage of the sum ofall precursors should be equal to the dosage range recited above forDHEA used alone, with appropriate adjustment for the different molecularweights of DHEA analogues such as DHEA esters. For example, if a DHEAester is used instead of DHEA, dosage should be increased by a multipleequivalent to the ratio of the molecular weight of the DHEA ester to themolecular weight of DHEA. The blood level of DHEA is the final criteriaof adequate dosage which takes into account individual variation inabsorption and metabolism.

Preferably, the attending clinician will, especially at the beginning oftreatment, monitor an individual patient's overall response and serumlevels of both estrogen and DHEA (in comparison to the preferred serumconcentrations discussed above), and monitor the patient's overallresponse to treatment, adjusting dosages as necessary where a givenpatients' metabolism or reaction to treatment is atypical. One approachwould be to start treating with DHEA or its analogue(s) alone and to addthe estrogen only if estrogen blood levels remain too low. Many patientsmay be treated only with the precursors of the invention withoutadditional estrogen.

Treatment in accordance with the invention is suitable for indefinitecontinuation. It is expected that DHEA treatment will simply maintainDHEA levels within a range similar to that which occurs naturally inwomen before menopause (serum concentration between 4 and 7 microgramsper liter), or naturally in young adult men (serum concentration between4 and 7 micrograms per liter). Accordingly, undesirable side effectsfrom sustained DHEA treatment are expected to be either minimal ornonexistent. Avoiding side effects from sustained estrogen use may beachieved in ways already known to the art, for example, by intermittentadministration of a progestin (e.g., medroxy-progesterone acetate) at adaily oral dose of 2 to 10 mg.

In order to facilitate the combination therapy of the invention, theinvention contemplates pharmaceutical compositions which include boththe estrogen and the second active compound (the precursor) in a singlecomposition for simultaneous administration. The composition may besuitable for administration in any traditional manner including but notlimited to oral administration, subcutaneous injection or intramuscularinjection. In other embodiments, a kit is provided wherein the kitincludes the estrogen and second compound (precursor(s)) in separatecontainers. In addition to other modes of administration, the secondcompound as well as the estrogen may also be administered transdermallyin accordance with the invention as discussed in more detail below.Thus, the kit may include appropriate materials for transdermaladministration, e.g., ointments, lotions, gels, creams, sustainedrelease patches and the like.

Applicants have discovered that administration of DHEA has utility inthe treatment of vaginal atrophy, hypogonadism and diminished libido,and improves the overall balance of circulating sex steroids, includingestrogens and androgens. It is believed that the prior art has notpreviously suggested that these conditions respond to DHEA treatment. Itis believed that DHEA, DHEA-S or a compound converted in vivo to eithermay be useful in the treatment of each of these three disorders.

Prior art methods for systemic administration of DHEA have included fororal and injection. Because DHEA treatments are often of prolonged andindefinite duration, repeated delivery by injection is veryinconvenient. Oral administration, however, has proven relativelyinefficient because orally administered DHEA goes first to the liverwhere a large percentage of it is prevented from entering the generalcirculation by local degradation.

We have recently observed that DHEA is very efficiently absorbedsystemically in both males and females following application to the skinor mucosa (e.g. buccal, vaginal or rectal mucosa). We have discoveredthat therapeutically efficient doses of DHEA may be administered by thepercutaneous or transmucosal route, thus avoiding first passage of thesteroid through the liver as results from oral administration, andfurther avoiding the discomfort and inconvenience of administering DHEAby injection.

Accordingly, the present invention provides delivery systems for theadministration of DHEA, DHEA-S and analogous compounds converted in vivoto either through the skin or mucosa. These systems are believed to bemore efficient than oral administration because the liver is bypassed.These systems are also significantly less painful and more convenientthan injections.

When DHEA, DHEA-S, or analogous compounds converted to DHEA or DHEA-S invivo are formulated for transdermal penetration, any of a number ofart-recognized transdermal penetration systems may be utilized. Forexample, DHEA may be prepared as part of an ointment, lotion, gel orcream for rubbing onto a patient's skin. Active ingredient is preferablypresent at from 7% to 20% by weight relative to the total weight of thepharmaceutical composition more preferably between 8 and 12%.Alternatively, the active ingredient may be placed into a transdermalpatch having structures known in the art, for example, structures suchas those set forth in E.P. Patent No. 0279982.

When formulated as an ointment, lotion, gel or cream or the like, theactive compound is admixed with a suitable carrier which is compatiblewith human skin or mucosa and which enhances transdermal penetration ofthe compound through the skin or mucosa. Suitable carriers are known inthe art and include but are not limited to Klucel HF and Glaxal base.Some are commercially available, e.g., Glaxal base available from GlaxalCanada Limited Company. Other suitable vehicles can be found in Kollerand Buri, S.T.P. Pharma 3(2), 115-124, 1987. The carrier is preferablyone in which the active ingredient(s) is (are) soluble at ambienttemperature at the concentration of active ingredient that is used. Thecarrier should have sufficient viscosity to maintain the precursor on alocalized area of skin or mucosa to which the composition has beenapplied, without running or evaporating for a time period sufficient topermit substantial penetration of the precursor through the localizedarea of skin or mucosa and into the bloodstream where it will cause ameasurable and desired increase in serum DHEA concentration. The carrieris typically a mixture of several components, e.g. pharmaceuticallyacceptable solvents and a thickening agent. A mixture of organic andinorganic solvents can aid hydrophylic and lipophylic solubility, e.g.water and an alcohol such as ethanol. Desirably, the carrier is onewhich, if formulated as 10% DHEA and 90% carrier (by weight) and appliedtwice daily in an amount providing 100 mg of DHEA to the abdominal area,will elevate serum concentration of DHEA in a typical patient by atleast 0.35 micrograms per liter per 50 kg of boby weight.

The carrier may also include various additives commonly used inointments and lotions and well known in the cosmetic and medical arts.For example, fiagrances, antioxidants, perfumes, gelling agents,thickening agents such as carboxymethylcellulose, surfactants,stabilizers, emollients, coloring agents and other similar agents may bepresent. When used to treat systemic diseases, the site of applicationon the skin should be changed in order to avoid excess localconcentration of steroids and possible overstimulation of the skin andsebaceous glands by androgenic metabolites of DHEA.

The precursor can also be administered, in some instances, by the oralroute, and may be formulated with conventional pharmaceuticalexcipients, e.g. spray dried lactose and magnesium stearate into tabletsor capsules for oral administration at concentrations providing easydosage in a range from 0.25 to 2.5 grams per day per 50 kg of bodyweight.

The active substance can be worked into tablets or dragee cores by beingmixed with solid, pulverulent carrier substances, such as sodiumcitrate, calcium carbonate or dicaldum phosphate, and binders such aspolyvinyl pyrrolidone, gelatin or cellulose derivatives, possibly byadding also lubricants such as magnesium stearate, sodium laurylsulfate, "Carbowax" or polyethylene glycol. Of course, taste-improvingsubstances can be added in the case of oral administration forms.

As further forms, one can use plug capsules, e.g. of hard gelatin, aswell as closed solf-gelatin capsules comprising a softner orplasticizer, e.g. glycerine. The plug capsules contain the activesubstance preferably in the form of granulate, e.g. in mixture withfillers, such as lactose, saccharose, mannitol, starches, such as potatostarch or amylopectin, cellulose derivatives or highly dispersed silicicacids. In solf-gelatin capsules, the active substance is preferablydissolved or suspended in suitable liquids, such as vegetable oils orliquid polyethylene glycols.

The concentration of active ingredient in the ointment, cream, gel orlotion is typically from about 7 to 20 percent preferably between 8 and12 percent and most preferably 10 percent (by weight relative to thetotal weight of the lotion, cream, gel or ointment). Within thepreferred ranges, higher concentrations allow a suitable dosage to beachieved while applying the lotion, ointment, gel or cream to a lessersurface area of the skin than would be possible at lower concentrationsand allows more freedom in choosing the body parts to which the ointmentor lotion will be applied. For example, it is well known in the art thata compound which is capable of transdermal penetration normallypenetrates more efficiently at some points in the body than in others.For example, penetration is very efficient on the forearm andconsiderably less effident on the palms.

The lotion, ointment, gel or cream should be thoroughly rubbed into theskin so that no excess is plainly visible, and the skin should not bewashed in that region until most of the transdermal penetration hasoccurred preferably at least 15 minutes and, more preferably, at least30 minutes.

A transdermal patch may be used to deliver precursor in accordance withknown techniques. It is typically applied for a much longer period,e.g., 1 to 4 days, but typically contacts active ingredient to a smallersurface area, allowing a slow and constant delivery of activeingredient.

A number of transdermal drug delivery systems that have been developed,and are in use, are suitable for delivering the active ingredient of thepresent invention. The rate of release is typically controlled by amatrix diffusion, or by passage of the active ingredient through acontrolling membrane.

Mechanical aspects of transdermat devices are well known in the rat, andare explained, for example, in U.S. Pat. Nos. 5,162,037, 5,154,922,5,135,480, 4,666,441, 4,624,665, 3,742,951, 3,797,444, 4,568,343,5,064,654, 5,071,644, 5,071,657, the disclosures of which areincorporated herein by reference. Additional background is provided byEuropean Patent 0279982 and British Patent Application 2185187.

The device may be any of the general types known in the art includingadhesive matrix and reservoir-type transdermal delivery devices. Thedevice may include drug-containing matrixes incorporating fibers whichabsorb the active ingredient and/or carrier. In a reservoir-type device,the reservoir may be defined by a polymer membrane impermeable to thecarrier and to the active ingredient.

In a transdermal device, the device itself maintains active ingredientin contact with the desired localized skin surface. In such a device,the viscosity of the carrier for active ingredient is of less concernthan with a cream or gel. A solvent system for a transdermal device mayinclude, for example, oleic add, linear alcohol factate and dipropyleneglycol, or other solvent systems known in the art. The active ingredientmay be dissolved or suspended in the carrier.

For attachment to the skin, a transdermal patch may be mounted on asurgical adhesive tape having a hole punched in the middle. The adhesiveis preferably covered by a release liner to protect it prior to use.Typical material suitable for release includes polyethylene andpolyethylene-coated paper, and preferably silicone-coated for ease ofremoval. For applying the device, the release liner is simply peeledaway and the adhesive attached to the patient's skin. In U.S. Pat. No.5,135,480, the disclosure of which is incorporated by reference, Bannonet al. describe an alternative device having a non-adhesive means forsecuring the device to the skin.

The target serum concentration of DHEA is the same, regardless ofwhether sex steroid precursor is being used as part of a combinationtherapy for treatment of menopause or is being used for the treatment ofskin deterioration, vaginal atrophy, hypogonadism or diminished libidoin accordance with the invention or for the treatment of a wide varietyof conditions related to decreased secretion of DHEA by the adrenals. Itis pointed out that dosage of DHEA, DHEA-S or any analog discussedherein can all be correlated to a target serum concentration of DHEAbecause all are converted in vivo, either directly or indirectly, intoDHEA.

The percutaneous or transmucosal delivery system of the invention mayalso be used as a novel and improved delivery system for the preventionand/or treatment of osteoporosis or other diseases which respondfavorably to treatment with DHEA. The desired target serum levels forthese latter purposes is also the same as indicated above.

DHEA used for percutaneous or transmucosal application can be in theform of the free alcohol or of one or more of its derivatives, evalerate, benzoate, acetate, enanthate and fatty ester derivatives. Thedelivery of DHEA or of its analogues through the skin is an acceptable,comfortable and noninvasive way of administering such compound. It alsoavoids gastrointestinal irritation and degradation of the compound andtoxicologlc problems due to first passage through the liver beforereaching the general circulation.

One method for preventing or inhibiting growth of breast and endometrialcarcinoma cells is activation of the androgen receptor with an effectivecompound having an affinity for the receptor site such that it binds tothe androgen receptor at low concentrations while not significantlyactivating other classes of steroid receptors linked to potential sideeffects.

Since DHEA is a natural source of androgens (Labrie, Mol. Cell.Endocrinol. 78: C113-C118, 1991) and the secretion of this compoundmarkedly decreases during aging, its replacement should have minimalunwanted side effects.

Percutaneous or transmucosal delivery of DHEA in accordance with theinvention thus provides a novel method for prevention and therapy ofdiseases responsive to activation of the androgen receptor, e.g. boneloss, obesity, breast cancer, endometrial cancer, hypogonadism, loss oflibido, loss of muscle mass, loss of energy, and other aging processes.The invention is also useful for many diseases wherein activation of theestrogen receptor will have beneficial effects, especially osteoporosisand vaginal atrophy.

EXAMPLES OF SOME PREFERRED DERIVATIVES

Derivatives of DHEA or DHEA-S in accordance with the present inventionmay be made by the following approaches:

Esterification of the 3βfunction into an ester which can be cleaved byesterase (the cleavage does not generate toxic substances) andtransformation of the 17-keto group into oxazolidine or thiazoliodineswhich are unstable in the body and regenerate a natural precursor.

Formation at position 3β of an α-acyloxyalkyl ether which can be cleavedby esterase into an unstable hemiacetal. The decomposition of thishemiacetal regenerates the natural precursor.

Without modification of the 3β function, transformation of the 17-ketogroup into oxazolidine or thiazoliodines which are unstable in the bodyand regenerate DHEA or DHEA-S.

The compounds disclosed below are expected to convert in vivo intodehydroepiandrosterone (DHEA) or dehydroepiandrosterone-sulfate ##STR7##wherein R³ is hydroxy or sulfate. wherein R^(e) is selected from thegroup consisting of hydrogen, benzyl, aryl, straight- or branched-alkyl,straight- or branched-alkenyl and straight- or branched-alkynyl.

wherein Z is oxygen or sulfur.

Some derivatives of DHEA being more lipophilic than DHEA itself can bestocked in skin fat and advantageously release DHEA slowly over time.

In some preferred compounds of the invention, the function at position 3is an ester of sulfuric acid (or salts thereof), formic acid , aceticacid, benzoic acid, butyric acid, decanoic acid, enanthic acid, furoicacid, heptanoic acid, isocaproic acid, undecanoic acid, undecylenicacid, palmitic acid, phenylpropionic acid, pivalic acid, propionic acid,valerie acid, carbonic acid (preferably ethylcarbonate orbenzylcarbonate).

In some preferred compounds of the invention, DHEA (or DHEA-S) ismodified by a 17-substituent of the following formula: ##STR8## whereinR'" is selected from the group consisting of hydrogen, benzyl, straight-or branched-alkyl and straight- or branched-alkenyl.

Certain preferred 3β-ester derivatives of DHEA are listed below:

    ______________________________________                                         ##STR9##                                                                     COMPOUNDS              X                                                      ______________________________________                                        dehydroepiandrosterone-3β-formate                                                               HCO                                                    dehydroepiandrosterone-3β-acetate                                                               CH.sub.3 CO                                            dehydroepiandrosterone-3β-propionate                                                            CH.sub.3 CH.sub.2 CO                                   dehydroepiandrosterone-3β-butyrate                                                              CH.sub.3 (CH.sub.2).sub.2 CO                           dehydroepiandrosterone-3β-valerate                                                              CH.sub.3 (CH.sub.2).sub.3 CO                           dehydroepiandrosterone-3β-pivalate                                                              (CH.sub.3).sub.3 CCO                                   dehydroepiandrosterone-3β-benzoate                                                              C.sub.6 H.sub.5 CO                                     dehydroepiandrosterone-3β-furoate                                                               C.sub.4 H.sub.3 OCO                                    dehydroepiandrosterone-3β-cypionate                                                             C.sub.5 H.sub.9 (CH.sub.2).sub.2 CO                    dehydroepiandrosterone-3β-lactate                                                               CH.sub.3 CHOHCO                                        dehydroepiandrosterone-3β-decanoate                                                             CH.sub.3 (CH.sub.2).sub.8 CO                           dehydroepiandrosterone-3β-undecanoate                                                           CH.sub.3 (CH.sub.2).sub.10 CO                          dehydroepiandrosterone-3β-palmitate                                                             CH.sub.3 (CH.sub.2).sub.14 CO                          dehydroepiandrosterone-3β-ethylcarbonate                                                        C.sub.2 H.sub.5 OCO                                    dehydroepiandrosterone-3β-benzylcarbonate                                                       C.sub.6 H.sub.5 CH.sub.2 OCO                           dehydroepiandrosterone-3β-isocaproate                                                           (CH.sub.3).sub.2 (CH.sub.2).sub.3 CO                   dehydroepiandrosterone-3β-undecylenate                                                          H.sub.2 CCH(CH.sub.2).sub.8 CO                         dehydroepiandrosterone-3β-enanthate                                                             CH.sub.3 (CH.sub.2).sub.5 CO                           dehydroepiandrosterone-3β-phenylpropionate                                                      C.sub.6 H.sub.5 (CH.sub.2).sub.2 CO                    3β-hydroxymethoxy-5-androsten-17-one acetate                                                    CH.sub.3 CO.sub.2 CH.sub.2                             3β-hydroxymethoxy-5-androsten-17-one                                                            CH.sub.3 (CH.sub.2).sub.8 CO.sub.2 CH.sub.2            decanoate                                                                     ______________________________________                                    

Some other preferred DHEA derivatives are listed below:

    ______________________________________                                         ##STR10##                                                                    COMPOUNDS                 R'"      Z                                          ______________________________________                                        3β-hydroxy-5-androstene-17-spiro(1',3'-thiazolidine-4'-                                            C.sub.2 H.sub.5                                                                        S                                          ethyl carboxylate)                                                            3β-hydroxy-5-androstene-17-spiro(1',3'-thiazolidine-4'-                                            C.sub.6 H.sub.5 CH.sub.2                                                               S                                          benzyl carboxylate)                                                           3β-hydroxy-5-androstene-17-spiro(1',3'-thiazolidine-4'-                                            C.sub.6 H.sub.13                                                                       S                                          hexyl carboxylate)                                                            3β-hydroxy-5-androstene-17-spiro(1',3'-oxazolidine-4'-                                             C.sub.2 H.sub.5                                                                        O                                          ethyl carboxylate)                                                            3β-hydroxy-5-androstene-17-spiro(1',3'-oxazolidine-4'-                                             C.sub.6 H.sub.5 CH.sub.2                                                               O                                          benzyl carboxylate)                                                           3β-hydroxy-5-androstene-17-spiro(1',3'-oxazolidine-4'-                                             C.sub.6 H.sub.13                                                                       O                                          hexyl carboxylate)                                                            ______________________________________                                    

Corresponding 17-substituted analogs of DHEA-S may also be used.

EXAMPLES OF SYNTHESIS Example 1

3β-formyloxy-5-androsiene-17-one

Following the procedure described by Ringold (H. J. Ringold, et al., J.Am. Chem. Soc. 78, 816, 1956), dehydroepiandrosterone (2.88 g, 10 mmol)dissolved in 85% formic acid (100 mL) is heated at 60° C. for 1 h. Aftercooling, the mixture is poured into iced water and after 16 h, crystalsare filtered and dried in vacuo.

Example 2

3β-acetoxy-5-androstene-17-one

Dehydroepiandrosterone (2.88 g, 10 mmol) is dissolved in a mixture (100ml) of anhydride acetic and pyridine (1:1 v/v) and left at roomtemperature for 16 h. The mixture is then poured carefully into icedwater and after 16 h, crystals are filtered and dried in vacuo.

Example 3

Dehydroepiandrosterone-3β-undecanoate

A solution of undecanoyl chloride (10.2 g, 50 mmol) in CH₂ Cl₂ (50 ml)was added to a mixture of 5-androsteno3β-ol-17-one (11.53 g, 40 mmol),Et₃ N (14 ml, 100 mmol) and dimethylaminopyridine (0.6 g, 5 mmol) in CH₂Cl₂ (150 ml) at 5° C. After addition, the mixture was stirred overnightat room temperature. The CH₂ Cl₂ layer was washed successively withwater, 2N HCl (twice), 5% K₂ CO₃ (twice), brine and dried. Removal ofthe solvent gave the crude product which was recrystallized with amixture of n-hexane:benzene to give the pure product (13.66 g; 75%),M.P. 84°-85° C.; ¹ H-NMR (CDCl₃); δ 0.83-0.86 (m, 6H, C₁₈ --CH₃ andCH₃); 1.03 (s, 3H, C₁₉ --CH₃); 4.58-4.61 (m, 1H, C₃ --H); 5.37 (d, 1H,vinyl, J=4.89 Hz). ¹³ C-NMR (CDCl₃) δ: 220.90, 179.25, 139.99, 121.78,73.38, 51.69, 50.14, 47.49, 38.12, 36.94, 36.72, 35.81, 34.68, 31.87,31.47, 31.42, 30.76, 29.53, 29.44, 29.27, 29.23, 29.09, 27.73, 25.03,22.66, 21.86, 20.31, 19.33, 14.09, 13.53.

Example 4

3β-acyloxy-5-androstene-17-one

The above esters at position 3β of dehydroepiandrosterone are preparedas follows:

Dehydroepiandrosterone (10 mmol) is dissolved in pyridine (50 mL) andadded to a solution of acyl chloride (prepared from the correspondingacid and oxalyl chloride) in the same solvant (50 mL).Dimethylaminopyridine (10%) is then added and the mixture is left atroom temperature for 16 h. The mixture is then carefully poured intoiced water and extracted with ethyl acetate. The organic phase is washedwith diluted HCl, water, saturated sodium bicarbonate and water, driedand evaporated to dryness to give the ester.

Example 5

3β-benzyloxycarbonyloxy-5-androsten-17-one

To a stirred solution of dehydroepiandrosterone (2.88 % 10 mmol) inmethylene chloride (100 mL) is added dropwise benzylchloroformate, overa period of 30 min following the known procedure (F. Reber and T.Reichstein, Helv. Chim. Acta, 28, 1164, 1945). After stirring for 3 h,the mixture is washed with water and evaporated to dryness. The residueis then dissolved in acetone and precipitated in iced water. After 16 h,crystals are filtered and dried in vacuo.

Example 6

3β-ethoxyoxycarbonyloxy-5-androsten-17-one

Same procedure as described in example 5 except that ethylchloroformateis used instead of benzylchloroformate.

Example 7

3β-hydroxy-5-androstene-17-spiro-2'-(1',3'-thiazolidine-4'-ethylcarboxylate)

Following the procedure described by Djerassi (C. Djerassi, N. Crossleyand M. A. Kielczewski, J. Org. Chem. 27, 1112, 1962),dehydroepiandrosterone (2.88 g, 10 mmol) is dissolved in anhydrousethanol, sodium acetate is added followed by L-cysteine ethyl esterhydrochloride (18 g, 100 mmol) and the mixture is heated overnight underan argon atmosphere. The reaction mixture is then evaported undervacuum. Methylene chloride is added to predpitate excess of L-cysteineethyl ester hydrochloride . The solution is then filtered and thefiltrate is washed twice with water, dried on magnesium sulfate,filtered and evaporated under vacuum. The residue is triturated withethanol to give crystals.

Example 8

3β-hydroxy-5-androstene-17-spiro-2'-(1', 3'-thiazolidine-4'-benzylcarboxylate)

Same procedure as described in example 7 except that L-cysteine benzylester hydrochloride is used instead of L-cysteine ethyl esterhydrochloride.

Example 9

3β-hydroxy-5-androstene-17-spiro-2'-(1',3'-thiazolidine-4'-alkylcarboxylate)

Same procedure as described in example 7 except that differentL-cysteine alkyl esters hydrochloride (e.g. L-cysteine hexyl esterhydrochloride) are used instead of L-cysteine ethyl ester hydrochloride.

Example 10

3β-hydroxy-5-androstene-17-spiro-2'-(1',3'-oxazolidine-4'-ethylcarboxylate)

The same procedure as described in the Example 7 is used except theoxazolidine derivative is prepared using serine ethyl esterhydrochloride instead of L-cysteine ethyl ester hydrochloride.

Example 11

3β-hydroxy-5-androstene-17-spiro-2'-(1',3'-oxazolidine-4'-benzylcarboxylate)

Same procedure as described in example 10 except that serine benzylester hydrochloride is used instead of serine ethyl ester hydrochloride.

Example 12

3β-hydroxy-5-androstene-17-spiro-2'-(1',3'-oxazolidine-4'-alkylcarboxylate)

Same procedure as described in example 10 except that different serinealkyl esters hydrochloride (e.g. serine hexyl ester hydrochloride) areused instead of serine ethyl ester hydrochloride.

Example 13

3β-hydroxymethoxy-5-androsten-17-one acetate

To a solution of dehydroepiandrosterone (2.88 g, 10 mmol) in THF (100mL) is added sodium hydride (11 mmol, 60% in oil) at room temperatureunder an argon atmosphere. When all the sodium hydride has reacted,chloromethyl acetate (prepared from acetyl chloride and formaldehyde (orderivative) using ZnCl₂ as catalyst) is added and the mixture is heatedfor a few hours. After cooling, the mixture is poured into water andextracted with ethyl acetate. The organic phase is then washed withwater, dried, filtered and evaporated to dryness to give the desiredcompound.

EXAMPLES OF PHARMACEUTICAL COMPOSITIONS

In one aspect, the present invention involves applying DHEA (or itsanalogues) formulation to the skin or other epithelial tissue for a timeperiod sufficient to permit sufficient penetration of the compound forsystemic or topical action, as desired. The composition may be appliedas a gel, a cream, an ointment, a lotion or the like or may involve useof a delivery system as described in U.S. Pat. Nos. 3,742,951, 3,797,494or 4,568,343. Devices as described in U.S. Pat. Nos. 5,064,654,5,071,644 or 5,071,657 can also be used to facilitate steroidabsorption.

All the pharmaceutical compositions of the present invention may containappropriate preservatives known in the art.

The following non-limiting examples describe the preparation of atypical cream, lotion, gel and ointment, respectively. In addition tothese vehicles, one skilled in the art may choose other vehicles inorder to adapt to specific dermatologic needs.

Example 14

A typical lotion contains (W/W) 10% DHEA, 15% propylene glycol and 70%ethanol and water 5%.

Example 15

A typical gel contains (W/W) 10% DHEA, 5% propylene glycol, 0.2%Carbomer 940 (available as Carbopol 940^(R) from B. F. Goodrich), 40%water, 0.2% triethanolamine, 2% PFG-12-Buteh-16 (available as Ucon^(R)fluid 50 from Union Carbide), 1% hydroxypropyl and 41.6% ethanol (95%ethanol-5% water).

Example 16

A typical ointment contains (W/W) 10% DHEA, 13% propylene glycol, 74%petrolatum, 2.9% glycerylmonostearate and 0.1% polylparaben.

Example 17

A typical cream contains (W/W) 10% DHEA, 0.2% propylparaben, 5% lanolinoil, 7.5% sesame oil, 5% cetyl alcohol, 2% glyceryl monostearate, 1%triethanolamine, 5% propylene glycol, 0.1% Carbomer 940^(R) and 64.2%water.

DHEA permeability can be enhanced by various techniques in order toreduce the dose used. Methods and compositions for enhancingpermeability of an active compound can be found, for example, in U.S.Pat. Nos. 5,051,260, 4,006,218, 3551,554, 3,472,931, 4,568,343,3,989,816 and 4,405,616.

Although the present invention has been described in relation toparticular embodiments thereof, many other variations and modificationsand other uses will become apparent to those skilled in the art. It ispreferred, therefore, that the present invention be limited not by thespecific disclosure herein, but only by the appended claims.

What is claimed is:
 1. A method for inhibiting the development of, ortreating, diminished libido comprising administering to a patient inneed of such treatment an amount of at least one sex steroid precursorselected from the group consisting of dehydroepiandrosterone,dehydroepiandrosterone sulfate, and compounds converted in vivo toeither of the foregoing, effective to maintain serum DHEA levels in saidpatient at between 4 and 13 micrograms per liter.
 2. The method of claim1 wherein said precursor is administered percutaneously ortransmucosally.
 3. The method of claim 1 wherein said precursor isadministered percutaneously or transmucosally by applying apharmaceutical composition containing said precursor at a concentrationof at least 7% by weight relative to the total pharmaceuticalcomposition.